The goal of the proposed project is to develop a method of inactivating genes only in mature B cells. This will make it possible to study the function of genes in mature B cells in vivo, even if the gene in question is required for animal viability and/or development of mature B cells. Also, as other cell types will not be affected, interpretations of the phenotypes observed should be more straightforward. Therefore, the first Specific Aim is to express the Cre recombinase only in mature B cells. The primary strategy for doing this is to insert Cre and an internal ribosome entry site into the 3' untranslated region of the delta heavy chain locus. As this locus is only expressed following VDJ recombination of the IgH locus, expression should be tightly restricted to B cells. Moreover, the delta heavy chain is not expressed during B cell development, but is expressed at high levels in every mature resting B cell, prior to antigenic Stimulation. Thus, Cre will be inserted by homologous recombination into the delta locus of embryonic stem cells and mice will be made from these cells. These mice will be crossed to beta-galactosidase reporter mice to directly assess the tissue and cell-type expression pattern of Cre recombinase by a functional assay. In the second Aim, the gene for the ubiquitously expressed signaling component Shc will be altered by introducing loxP sites on either side of essential exons. This should not affect Shc expression except in cells expressing Cre, which will delete the DNA between the two loxP sites, inactivating the Shc gene. Thus, Shc should be lost in mature B cells, but should be normally expressed in all other cell types. The effects of this defect on mature B cell function will be assessed after the funding period.